Breathing is a vital, multifaceted behavior in which a centrally generated rhythm must be transformed into coordinated patterns of motor activity on spinal and cranial nerves that produce the pumping action of the chestwall and regulate airway tone. Mechanosensory input from the airways is essential for this coordination. Especially in mammals that have highly compliant chest walls - including human infants - antagonistic sets of airway receptors regulate end-inspiratory volume (ie, tidal volume) and end-expiratory volume. However, little is known about how these differential respiratory motor responses are coordinated within the central nervous system. In this application we will define (in rats) the central pathways by which two antagonistic reflexes are elicited in response to lung inflation (slowly adapting receptors;SARs) and lung deflation (deflation activated receptors;DARs). Three Specific Aims will be addressed. In Aim 1 the axonal projections of the different types of SAR and DAR NTS relay neurons will be comprehensively surveyed using anterograde labeling of small groups of NTS cells combined with retrograde labeling of their synaptic targets. Complementary studies using cross-correlation of simultaneously recorded spike trains will assess these functional interactions. In Aim 2, juxtacellular dye filling of single, uniquely defined SAR and DAR interneurons along with retrograde labeling of their targets will permit combined light- and electron-microscopic analysis of the synaptic connections of NTS relay neurons with multiple targets. In Aim 3 the electrophysiologically defined phenotypes of neurons post- synaptic to SAR and DAR NTS interneurons will be defined using juxtacellular labeling of both NTS neurons and their potential targets. Spike-triggered averaging will permit characterization of the synaptic connectivity, and its relative strength, between the NTS relay neurons and specific post-synaptic cells. Taken together, these experiments will identify, for the first time, the post-synaptic targets of SAR and DAR afferent relay neurons and provide a polysynaptic circuit diagram of the pathways producing coordinated respiratory reflex responses to activation of lung mechanoreceptors. Knowledge of these circuits may elucidate treatment strategies for patients with compromised airway control characteristic of a range of respiratory disorders and degenerative motor diseases. PROJECT NARRATIVE: This research will examine the brainstem circuits related to central reflexes modulating breathing. By following in detail the interconnections of neurons receiving information from the airways and lungs we will describe the control system for the force and temporal pattern of breathing in a way that has not previously been possible. This type of information will be important for analyzing the role of these circuits in a variety of human diseases ranging from amyotrophic lateral sclerosis to sudden infant death syndrome.